A general SNP-based molecular barcode for Plasmodium falciparum identification and tracking
A general SNP-based molecular barcode for Plasmodium falciparum identification and tracking
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DOI：
10.1186/1475-2875-7-223
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发表时间：
2008-10-29
期刊：
影响因子：
3
通讯作者：
Wiegand, Roger C.
中科院分区：
文献类型：
--
作者：
Daniels, Rachel;Volkman, Sarah K.;Wiegand, Roger C.
背景资料：单核苷酸多态性（SNP）基因分型提供了一种方法来开发一个实用的，快速的，廉价的检测，将唯一地确定任何恶性疟原虫寄生虫使用少量的DNA。这种检测可用于区分药物试验中的复发与再感染，监测接受药物治疗或疫苗挑战的患者群体中特定寄生虫的频率和分布，或用于在培养适应和亚克隆以及常规传代期间跟踪样品并确定实验室分离株的纯度。方法：已经鉴定了一组24个SNP标记，其表现出高的次要等位基因频率（平均MAF > 35%），针对其构建了稳健的TaqMan基因分型测定。通过全基因组测序鉴定所有SNP，并通过全球寄生虫收集的基于Affyphidae阵列的基因分型估计MAF。这些检测创建了一个“分子条形码”，以唯一地识别寄生虫genome.Results：使用24个这样的标记，没有两个寄生虫已知的独立起源尚未被发现有相同的等位基因签名。TaqMan基因分型检测可以在各种样品上进行，包括培养的寄生虫、冷冻的全血或滤纸上点样的全血，成功率> 99%。需要少于5 ng的寄生虫DNA来完成一组24个标记。这个SNP面板检测和识别寄生虫的能力进行了比较，以标准的分子方法，MSP-1和MSP-2 typing.Conclusion：这项工作提供了一个简单的现场部署的基因分型工具，可以使用没有特殊技能的标准实验室设备，并在合理的成本，将明确识别和跟踪恶性疟原虫寄生虫从患者样本和在实验室。
Background: Single nucleotide polymorphism (SNP) genotyping provides the means to develop a practical, rapid, inexpensive assay that will uniquely identify any Plasmodium falciparum parasite using a small amount of DNA. Such an assay could be used to distinguish recrudescence from re-infection in drug trials, to monitor the frequency and distribution of specific parasites in a patient population undergoing drug treatment or vaccine challenge, or for tracking samples and determining purity of isolates in the laboratory during culture adaptation and sub-cloning, as well as routine passage.Methods: A panel of twenty-four SNP markers has been identified that exhibit a high minor allele frequency (average MAF > 35%), for which robust TaqMan genotyping assays were constructed. All SNPs were identified through whole genome sequencing and MAF was estimated through Affymetrix array-based genotyping of a worldwide collection of parasites. These assays create a "molecular barcode" to uniquely identify a parasite genome.Results: Using 24 such markers no two parasites known to be of independent origin have yet been found to have the same allele signature. The TaqMan genotyping assays can be performed on a variety of samples including cultured parasites, frozen whole blood, or whole blood spotted onto filter paper with a success rate > 99%. Less than 5 ng of parasite DNA is needed to complete a panel of 24 markers. The ability of this SNP panel to detect and identify parasites was compared to the standard molecular methods, MSP-1 and MSP-2 typing.Conclusion: This work provides a facile field-deployable genotyping tool that can be used without special skills with standard lab equipment, and at reasonable cost that will unambiguously identify and track P. falciparum parasites both from patient samples and in the laboratory.